The present invention relates to macrolides with anti-inflammatory activity and, more particularly, it relates to des-dimethylamino macrolide derivatives with anti-inflammatory activity, their pharmaceutically acceptable salts and pharmaceutical compositions containing them as active ingredients.
It is known that many antibiotics, in particular the class of macrolides with 14 atoms derived from erythromycin, are endowed with anti-inflammatory properties in addition to the antibacterial activity [Clin. Immunother., (1996), 6, 454-464].
Erythromycin is a natural macrolide (The Merck Index, XII edition, nxc2x0 3720, page 625) that has had a very broad clinical use in the treatment of infections caused by Gram-positive bacteria, by some Gram-negative ones or by Mycoplasma.
Recently the interest of the scientific community has been focused on the anti-inflammatory and immunomodulatory component of erythromycin and derivatives [Journal of Antimicrobial Chemotherapy, (1998), 41, Suppl.B, 37-46].
Such activity is well documented by both clinical studies and in vivo and in vitro experiments.
For example, macrolides have proved to be effective in the therapy of inflammatory diseases such as panbronchiolitis [Thorax, (1997), 52, 915-918], bronchial asthma [Chest, (1991), 99, 670-673] and cystic fibrosis [The Lancet, (1998), 351, 420], or in animal models of inflammation such as, for example, the zymosan-induced peritonitis in mice [Journal of Antimicrobial Chemotherapy, (1992), 30, 339-348] and the neutrophil recruitment induced by endotoxin in rat trachea [Antimicrobial Agents and Chemotherapy, (1994), 38, 1641-1643] or in in vitro studies on immune system cells, such as neutrophils [The journal of Immunology, (1997), 159, 3395-4005] and T-lymphocytes [Life Sciences, (1992), 51, PL 231-236] or in the modulation of cytokines, such as interleukin 8 (IL-8) [Am. J. Respir. Crit. Care Med., (1997), 156, 266-271] or interleukin 5 (IL-5) (EP 0 775 489 and EP 0 771 564, Taisho Pharmaceutical Co., Ltd).
The peculiar therapeutic efficacy of macrolides on diseases in which the conventional anti-inflammatory drugs, such as for example corticosteroids, have demonstrated to be ineffective [Thorax, (1997), 52, 915-918, already cited] justifies the high interest towards this new potential class of anti-inflammatories.
Nevertheless the strong antibacterial activity of the conventional macrolides does not allow an enlarged use in the chronic treatment of inflammatory processes not due to pathogens because of the rapid onset of resistant strains.
Therefore, it would be desirable to have new substances with a macrolide structure that show anti-inflammatory activities and, in the mean time, that are devoid of antibiotic properties.
For a greater clarity we show the formula of erythromycin in which the numbering adopted in the present patent application is indicated. 
Some classes of erythromycin derivatives endowed with a high anti-inflammatory activity are described in the literature.
For example, in the already cited European patent applications in the name of Taisho derivatives of erythromycin modified at 3, 9, 11 and 12 position, as strong inhibitors of the synthesis of IL-5 are claimed.
N-alkyl derivatives of azithromycin, without cladinose and desosamine, of formula 
wherein
R1 is hydrogen, a lower alkyl or a lower alkanoyl; R2, R3 and R4, the same or different, are hydrogen or a lower alkanoyl; are described as anti-inflammatories in EP 0 283 055 (Sour Pliva).
The use of erythromycin as anti-inflammatory that acts by reducing the release of interleukin 1 through the inhibition of the mammalian mdr-P glycoprotein is claimed WO 92/16226 in the name of Smith-Kline Beecham Corporation.
Among the macrolide derivatives described in the literature a few are 3xe2x80x2-desdimethylamino-9-oxyimino derivatives. The limited interest towards this class of compounds is justified by the fact that the relevance of the dimethylamino group for the activity of ribosomal binding typical of macrolides is known [Tetrahedron Letters, (1994), 35, 3837-3840].
In U.S. Pat. No. 3,928,387 (Hoffmann-La Roche Inc.) 3xe2x80x2-desdimethylamino-3xe2x80x2,4xe2x80x2-dehydroerythromycin A oxime is described, as intermediate useful for the preparation of the antibiotic 1745A/X.
In EP 0 254 534 (Robinson, William S.) a very broad class of macrolides with antiviral activity is claimed. Among them the compound of formula 
whose correct chemical name is 3xe2x80x2-desdimethylamino-3xe2x80x2,4xe2x80x2-dehydroerythromycin A 9-O-methyloxime notwithstanding in the text of EP 0 254 534 is erroneously reported as des-dimethylaminoerythromycin 9-O-methyloxime (page 10, line 46) is described.
Now we have found that by removing the dimethylamino group from the 3xe2x80x2 position of desosamine of 9-oxyimino macrolides, compounds endowed with anti-inflammatory activity and essentially devoid of antibiotic properties are obtained.
Therefore, object of the present invention are compounds of formula 
wherein
R is hydrogen or methyl;
R1 and R2 are both hydrogen or they together form a bond;
R3 is hydrogen, a linear or branched C1-C5 alkyl group, a benzyl group, optionally substituted by one or more substituents selected among nitro groups, hydroxy groups, carboxylic groups, amino groups, linear or branched C1-C5 alkyl groups, C1-C4 alkoxycarbonyl groups, aminocarbonyl groups or ciano groups, or a chain of formula 
xe2x80x83wherein
A is hydrogen or a phenyl group optionally substituted by one or two substituents selected among nitro groups, hydroxy groups, carboxylic groups, amino groups, linear or branched C1-C5 alkyl groups, C1-C4 alkoxycarbonyl groups, aminocarbonyl groups or ciano groups, or a 5 or 6 membered heterocycle, saturated or unsaturated, containing from 1 to 3 heteroatoms selected among nitrogen, oxygen and sulphur, optionally substituted by one or two substituents selected among C1-C5 alkyl groups, phenyl groups, hydroxy groups, oxo (xe2x95x90O) groups, nitro groups, C1-C4 alkoxycarbonyl groups, aminocarbonyl groups, mono or di-C1-C4-alkylaminocarbonyl groups, C1-C4-alkylcarbonyl groups;
X and Y, the same or different, are O, S, SO, SO2 or NR4, in which R4 is hydrogen, a linear or branched C1-C5 alkyl group, a C1-C5 alkoxycarbonyl group, a benzyloxycarbonyl group;
r is an integer from 1 to 6;
m is an integer from 1 to 8;
n is an integer from 0 to 2;
and their pharmaceutically acceptable salts; the compounds 3xe2x80x2-desdimethylamino-3xe2x80x2,4xe2x80x2-dehydroerythromycin A oxime (R1 and R2=bond; Rxe2x95x90H; R3xe2x95x90H) and 3xe2x80x2-desdimethylamino-3xe2x80x2,4xe2x80x2-dehydroerythromycin A 9-O-methyloxime (R1 and R2=bond; Rxe2x95x90H; R3xe2x95x90CH3) being excluded.
A further object of the present invention is the use of the compounds 3xe2x80x2-desdimethylamino-3xe2x80x2,4xe2x80x2-dehydroerythromycin A oxime and 3xe2x80x2-desdimethylamino-3xe2x80x2,4xe2x80x2-dehydroerythromycin A 9-O-methyloxime as anti-inflammatories.
The compounds of formula I are anti-inflammatory macrolides devoid of antibiotic activity and therefore they are useful in the treatment of inflammatory diseases.
With the term linear or branched C1-C5 alkyl groups a group selected among methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl and isopentyl is meant. With the term 5 or 6 membered heterocycle, saturated or unsaturated, containing from 1 to 3 heteroatoms selected among nitrogen, oxygen and sulphur, heterocycles such as pyrrole, thiophene, furan, imidazole, pyrazole, thiazole, isothiazole, isoxazole, oxazole, pyridine, pyrazine, pyrimidine, pyridazine, triazole, thiadiazole and their partially o totally saturated forms are meant.
Preferred compounds of formula I are the compounds in which R, R1 and R2 are hydrogen.
Within this class the compounds in which R3 is a chain of formula 
wherein
X, Y, A, r, m and n have the already reported meanings are particularly preferred.
Even more preferred compounds are the compounds in which R3 is a chain of formula 
wherein
r is 2, m is 2 or 6, n is 1, Y is NR4, X is O or NR4, R4 is hydrogen and A is phenyl or thiazolyl.
Examples of pharmaceutically acceptable salts of the compounds of formula (I) are the salts with organic or inorganic acids such as hydrochloric, hydrobromic, hydriodic, nitric, sulphuric, phosphoric, acetic, tartaric, citric, benzoic, succinic and glutaric acid.
The compounds of formula I, object of the present invention, are prepared by following a synthetic scheme which comprises (a) the removal of the dimethylamino group at 3xe2x80x2 position and (b) the optional functionalisation of the oxime.
The removal of the dimethylamino group is performed by oxidation, pyrolisis and optional reduction, according to known methods. It is evident to the man skilled in the art that, in order to avoid interferences with the functional groups optionally present at R3 substituent, the removal of the dimethylamino group will be preferably accomplished starting from intermediates of formula 
in which
R has the already reported meanings and R3xe2x80x2 is hydrogen or a linear or branched C1-C5 alkyl group.
By oxidation the corresponding N-oxides of formula 
in which
R and R3xe2x80x2 have the already reported meanings are obtained; that by pyrolisis, optionally followed by reduction, respectively give the compounds of formula 
object of the present invention (I-R3=hydrogen or C1-C4 alkyl).
According to common techniques, the compounds of formula I in which R3 is different from hydrogen can be prepared from the compounds of formula I-A and I-B in which R3xe2x80x2 is hydrogen by functionalisation of the oxime. Generally the functionalisation is carried out by reaction with a compound of formula
R3xe2x80x3xe2x80x94Wxe2x80x83xe2x80x83(IV)
in which R3xe2x80x3 has all the meanings of R3 apart from hydrogen and W is a leaving group, preferably a chlorine or bromine atom or a mesyl group.
An alternative synthetic route particularly suited for the preparation of the compounds of formula I in which R3 is a chain of formula 
in which
X, Y, A, r, m and n have the already reported meanings;
comprises the reaction of a compound of formula I in which R3 is hydrogen with an intermediate of formula 
in which
W, X, Y, m and n have the already reported meanings and Z represents a protecting group;
to give the intermediate of formula 
in which R, R1, R2, X, Y, Z, r and m have the already reported meanings;
that after removal of the Z protecting group, is reacted with a derivative of formula 
in which A, W and n have the already reported meanings;
to give the compounds of formula I.
The compounds of formula I in which Y is NR4 can be prepared according to the above reported synthetic route also by using an aldehyde of formula
xe2x80x83Axe2x80x94CHOxe2x80x83xe2x80x83(VIII)
in which A has the already reported meanings;
in place of the intermediate of formula VII, subject to removal of the Z protecting group from the intermediate of formula VI.
Moreover, the compounds of formula I in which R1xe2x95x90R2xe2x95x90H can be prepared by reduction of the corresponding compounds of formula I in which R1 and R2 form a bond.
The compounds of formula I, object of the present invention, are endowed with an anti-inflammatory activity and are devoid of antibiotic activity.
The pharmacological activity of the compounds of formula I has been evaluated by in vitro and in vivo tests in comparison with known macrolides, such as erythromycin, clarithromycin and roxithromycin, endowed with both anti-inflammatory activity and antibiotic activity.
The anti-inflammatory activity has been evaluated in vitro as inhibition of IL-8 release and of superoxide anion release (example 11) and in vivo as inhibition of LPS-induced neutrophilia after repeated administrations (example 12).
In all the experiments the compounds object of the present invention have resulted very active as anti-inflammatories and the anti-inflammatory activity has been equal to or greater than the one of the reference compounds.
For a therapeutical application the compound of formula I can be used in a pharmaceutical form suitable to oral or parenteral administration.
Therefore, object of the present invention are pharmaceutical compositions containing a therapeutically active amount of a compound of formula I or of a salt thereof in admixture with a pharmaceutically acceptable carrier.